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目的:建立一种快速明确饮用水中是否存在细菌污染的核酸检测方法。方法:根据细菌16SrRNA中高度保守的序列设计4条LAMP引物,交由商业公司合成;利用合成好的引物和标准菌株,通过LAMP方法验证引物扩增的效率和特异性,电泳观察扩增结果,并比较两种不同细菌基因组DNA提取方法及优化LAMP反应条件。结果:使用设计的引物可以很好的检测出各种标准菌株,而且检测灵敏度达到要求。使用碱裂解的方法可以非常方便的提取出细菌基因组DNA。结论:成功建立了LAMP法检测饮用水中细菌的方法,为饮用水是否存在细菌污染快速检测提供了一种可用的方法。
Objective: To establish a rapid and clear determination of the presence of bacterial contamination in drinking water nucleic acid detection methods. METHODS: Four LAMP primers were designed according to the highly conserved sequences in bacterial 16S rRNA and were synthesized by commercial companies. The efficiency and specificity of primer amplification were verified by LAMP assay using synthetic primers and standard strains. The amplification results were observed by electrophoresis. The genomic DNA extraction methods from two different bacteria were compared and the LAMP reaction conditions were optimized. Results: Using the designed primers can detect a variety of standard strains well, and the detection sensitivity meets the requirements. Bacterial genomic DNA can be very conveniently extracted by alkaline lysis. Conclusion: The LAMP method for the determination of bacteria in drinking water has been successfully established, which provides a useful method for rapid detection of bacterial contamination in drinking water.